Stem cells are a key focus in tissue engineering strategies for clinical applications. However, the ability to inject and localize them to a target site while maintaining predictably high cell viability is critically lacking. We have shown that the viscoelastic properties of an injection matrix in part determine the relative viability of a variety of different stem cells after injection. When suspended in PBS, the simple act of injection through a 28 gauge syringe needle leads to compromised cell viability. When cells are instead suspended within a shear-thinning alginate hydrogel, ejected cell viability of human umbilical vein endothelials cells (HUVECs), human adipocyte stem cells (hASCs), rat mesenchymal stem cells (rMSC), and murine neural stem cells (mNSC) can be improved by 10-30%.
Common shear-thinning, reversibly gelling matrices such as collagen and Matrigel utilize non-physiological environmental triggers (e.g., pH and temperature shifts) detrimental to cells. We have recently developed a mixture-induced two component hydrogel (MITCH) that undergoes gelation under constant physiological conditions. The two components are both recombinantly-engineered protein polymers that include either the WW domain or its proline-rich ligand. These two domains hetero-assemble upon mixing through simple hydrogen bonding. Microrheology experiments confirm gelation within one minute after mixing to produce moduli ranging from ~10 Pa to ~50 Pa, dependent on the choice of molecular recognition binding partners. These gels undergo rapid shear-thinning and can re-form within 5 minutes, exemplifying their self-healing nature. This physically gelling material allows biocompatible cell encapsulation; for example, hASC viability immediately post-encapsulation is 97 ± 2%. MITCH mechanical properties are also suitable for neural encapsulation, and mNSCs encapsulated within MITCH remain highly viable and proliferative. Encapsulated and differentiated mNSCs stain positive for Map2 and GFAP, indicating neuronal and glial cells, respectively. These cells adopt 3D-branched morphologies with elongated dendrites extending over 100 μm.
Here we show that the viscosity of an injection material strongly influences the viability of cells ejected through a clinically relevant syringe needle. Additionally, MITCH materials promote the differentiation of neural progenitors into neuronal phenotypes, which adopt a 3D-branched morphology within the hydrogels. The modular hetero-assembly of MITCH materials also permits the tethering of neuroprotective peptides to the gel, facilitating further optimization for the prevention and treatment of central nervous system injuries.